Method and system for replacing the water cooled laser in a microplate reader

ABSTRACT

The present invention relates to a method and system for repairing and refurbishing a microplate reader of the Flipr type which has a water cooled argon laser light source. The old laser is removed and replaced with a high power (300 to 500 mW) air cooled solid state laser as a replacement place on its own support and focused and wired to replace the old laser. The new product operates at lower power consumption yet provides accurate measurements.

This application claims priority of U.S. provisional application61/147,080 filed on Jan. 24, 2009 and is included herein in its entiretyby reference.

COPYRIGHT NOTICE

A portion of the disclosure of this patent contains material that issubject to copyright protection. The copyright owner has no objection tothe reproduction by anyone of the patent document or the patentdisclosure as it appears in the Patent and Trademark Office patent filesor records, but otherwise reserves all copyright rights whatsoever.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to a method and system for replacing alighting system in an existing microplate reader with a new lightsource. Specifically, the present invention relates to a system whichcan be used to replace a water cooled laser with an LED light system inan existing microplate reader.

2. Description of Related Art

The use of fluorescence analytical monitoring techniques is well known.Fluorescence measurements can be taken by shining a light source of afirst wavelength and then when the sample of light is absorbed, the testmaterial is induced to emit light of a second wavelength. Measurement ofthe second wavelength light, either as length, intensity or the like,can be used to correlate the activity in a cell that is producing thesecondary fluorescence. Physiological parameters can then be determinedbased on the results, such as potassium or other ion channel activity.Florescence type measurements are of great importance to the researchand development of new pharmaceutical compositions and are used toscreen a variety of tissues for interaction with most any chemicalcomposition that is of interest in affecting the measurable systems.

Typically, in the analysis of cells, a variety of older machines areavailable for using this type of technique with 96 or 384 well multiplewell plates. These machines provide a multiwall plate holder, a watercooled laser light source and some form of a receiving camera fordetecting the cell's second wavelength light emissions. The cells arecultured in each of the wells at the bottom with a growth mediumprovided over the growing cells. The chemical compound to be tested orotherwise assayed is played into the liquid in each well with theflorescent material and the effect measured by excitation by the laserand reading by the camera. For example, in U.S. Pat. No. 5,355,215 toSchroder et al issued Oct. 11, 1994, there is disclosed a method foraligning a camera and a light source for measuring the second wavelengthwith a minimum about if interference from the supernate liquid. Thisprocess has been utilized extensively and appears in later machines andin more recently issued patents, for example, in U.S. Pat. No. 7,265,829issued Sep. 4, 2007 to Jiang et al.

The older microplate reader machines that utilize water cooled lasers,while difficult to use, were well built and very cost effective. Suchmachines include the Flipr2® and Flipr3® microplate readers. Thesemachines suffered from the difficulty of using and operating watercooled lasers but because parts were relatively accessible for repair, aburgeoning business in repair and refurbishment developed to keep thesemachines in service. Since the refurbishment of even a patented productis allowable repair, such repair has been accomplished not only by OEMs,but a variety of small companies also repair these types of machines.The lasers in these machines are typically in the 3 watt water cooledargon laser with power designed to be sufficient in order to providesufficient light to produce excitation of each well in a microplate.

Newer microplate reader machines are very costly and tend to be large inan attempt to avoid the difficulties in using water cooled lasers. Anexample is the Flipr^(tetra)® made by the same company as the olderFlipr2 and Flipr3 machines, Molecular Devices (MDC). In order toencourage purchases of newer models of microplate readers, MDC hasdeclined to support its older machines and encourage users of the oldermachines to upgrade. However, for many users the older machines are fineand there is a desire in the marketplace to continue refurbishing thesemachines. The fact that water cooled lasers that these machine werebuilt to utilize are expensive, hard to use and getting scarce to fitthe existing machines, has suggested the repair life of these machinesis limited in spite of the desire of users to continue using themachines to the end of their useful life.

Digital lasers are now well known and can be obtained in a variety ofwattages. However, the wattages of the air cooled laser types has notyet reached the 3 watts that are currently employed and rated for theexisting machines with water cooled lasers. In fact, currently thelargest air cooled lasers are in the 500 mW range and industriallyconsidered not powerful enough to be used in a microplate reader.

BRIEF SUMMARY OF THE INVENTION

The use of argon water cooled lasers has been extensively used bymicroplate readers. These lasers are sensitive and easily misaligned.The microplate readers used with them were designed, shaped andcalibrated specifically to be used with these types of lights sources.It has been discovered that it is possible to replace the light sourcewith 300 to 500 mW air cooled laser light source under specificconditions and repair such a unit and use filters holders supports, andthe like, of the present invention thus extending the useful repair lifeof these older microplate readers without the problems normallyassociated with argon water cooled lasers and surprisingly with a laserone-sixth the power of OEM lasers in microplate readers with watercooled lasers.

In one embodiment of the present invention, there is a method forrepairing a fluorescence microplate reader having a water cooled laserlight source of about 3 watts that has been removed from the readercomprising:

-   -   a) selecting an air cooled laser having a wattage of between        about 300 and 500 mW, having a total lumen output sufficient to        produce a replacement lumen output, the air cooled laser mounted        in a rigid support and provide light to the microplate reader;    -   b) providing that the light output of the air cooled laser is at        a desired wavelength;    -   c) positioning the support such that the light from the air        cooled laser shines on the bottom of a microplate positioned in        the reader; and    -   d) connecting the air cooled laser to the reader such that they        operate in place of the water cooled laser light.

In another embodiment of the invention, there is a system for replacinga water cooled laser in a fluorescence microplate reader comprising:

-   -   a) An air cooled laser having a power of about 300 to 500 mW        mounted in fixed relationship having a lumen output sufficient        to produce a replacement lumen output;    -   b) an optional filter for changing the wavelength of the LED        lights to a desired wavelength where the wavelength of the air        cooled laser is not originally a desired wavelength;    -   c) a support for adjustably positioning the air cooled laser and        optionally the filter in the reader such that when placed in the        reader the light from the air cooled laser shine on the bottom        of a microplate placed in the reader.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a side view of a microplate reader with an air cooled laserlight source positioned partially outside the reader.

FIG. 2 is a side view of a microplate reader with an air cooled laserlight source positioned inside the reader and using a dichroic mirror.

DETAILED DESCRIPTION OF THE INVENTION

While this invention is susceptible to embodiment in many differentforms, there is shown in the drawings and will herein be described indetail specific embodiments, with the understanding that the presentdisclosure of such embodiments is to be considered as an example of theprinciples and not intended to limit the invention to the specificembodiments shown and described. In the description below, likereference numerals are used to describe the same, similar orcorresponding parts in the several views of the drawings. This detaileddescription defines the meaning of the terms used herein andspecifically describes embodiments in order for those skilled in the artto practice the invention.

The terms “a” or “an”, as used herein, are defined as one or as morethan one. The term “plurality”, as used herein, is defined as two or asmore than two. The term “another”, as used herein, is defined as atleast a second or more. The terms “including” and/or “having”, as usedherein, are defined as comprising (i.e., open language). The term“coupled”, as used herein, is defined as connected, although notnecessarily directly, and not necessarily mechanically.

Reference throughout this document to “one embodiment”, “certainembodiments”, and “an embodiment” or similar terms means that aparticular feature, structure, or characteristic described in connectionwith the embodiment is included in at least one embodiment of thepresent invention. Thus, the appearances of such phrases or in variousplaces throughout this specification are not necessarily all referringto the same embodiment. Furthermore, the particular features,structures, or characteristics may be combined in any suitable manner inone or more embodiments without limitation.

The term “or” as used herein, is to be interpreted as an inclusive ormeaning any one or any combination. Therefore, “A, B or C” means any ofthe following: “A; B; C; A and B; A and C; B and C; A, B and C”. Anexception to this definition will occur only when a combination ofelements, functions, steps or acts are in some way inherently mutuallyexclusive.

The drawings featured in the figures are for the purpose of illustratingcertain convenient embodiments of the present invention, and are not tobe considered as limitation thereto. Term “means” preceding a presentparticiple of an operation indicates a desired function for which thereis one or more embodiments, i.e., one or more methods, devices, orapparatuses for achieving the desired function and that one skilled inthe art could select from these or their equivalent in view of thedisclosure herein and use of the term “means” is not intended to belimiting.

As used herein a “fluorescence microplate reader” refers to thosemicroplate readers which originally were designed to handle 384 platereaders or less and use a water cooled laser such as an argon watercooled laser to provide an excitation light source. Argon lasers providea light of a wavelength of about 488 nanometers. Units such as theFLIPR2 and FLIPR3 unit made by MDC are examples of the intendedmicroplate readers that are repaired using the method and system of thepresent invention. While the argon lasers do work for their intendedpurpose in these types of microplate readers, they are difficult to use,have a long warm up time and need to be frequently adjusted. Whenreplacing the lasers during the repair or refurbishment process, it isunderstood that the laser would be removed before beginning with themethod and system of the present invention. These types of microplatereaders have an upper portion where pipetting is done to a microplatesitting or placed in the reader. Once a microplate is placed in thereader the bottom surface of the microplate is exposed to the bottomportion of the reader where a light source, such as the argon laser, canshine on the bottom surface of the plate specifically so that it shineson each well bottom surface. Light emissions from the microplate wellsoccurs in the lower position as well and reaches a CCD type cameraeither directly or indirectly by any of one or more mirrors.

As used herein, “air cooled laser” refers to digital air cooled lasers.These are also referred to as optically pumped semiconductor lasers.These are the highest power air cooled solid state lasers currentlycommercially available. They are available in wattages of about 500 mWand below and have a wavelength light of 488 nm. These are consideredhigh power lasers of these types even though much higher power lasersare available in the argon water cooled typed which came stock in thetype of microplate readers being repaired by the method of thisinvention. These are typically a blue light laser and certainly couldhave a range of blue light from about 425 to about 510 nm. One exampleof such a unit is the Sapphire 488-500 Laser head produced as a 500 mWCW blue laser having an output of 488 nm by Coherent. It is known in theart that these type of lasers work with their own controller andfrequently an appropriate heat-sink instead of the water cooled versionsof lasers. One skilled in the art in view of this disclosure could adaptthe air cooled laser accordingly. While the unit being replaced has avery high power 3 watt laser. It has been surprisingly found thatreplacing the water cooled laser with an air cooled high power laser offrom about 300 mW up to the maximum 500 mW actually surprisingly stillworks as a light source for a microplate reader when replacing it fromthe standard water cooled laser.

It has also been discovered that a more homogeneous lighting can beachieved by filtering the light or passing the light from the air cooledlaser through a filter or aperture of some kind. These embodimentsprovide a more even lighting on the bottom of a microplate duringreading, and thus when tested, results in a coefficient of variation(CV) of 0.6% or about the same as a water cooled laser. One skilled inthe art and understanding the problems described herein can vary thechoice of air cooled lasers and filters or apertures to optimize thelight source in view of the disclosure herein.

The air cooled laser needs to be held in relative fixed relationship sothat light emanating from the air cooled laser remains constant withinthe beam produced. The stand or mounting bracket, or the like, can keepthe relative position of the air cooled laser constant but must be ableto be adjusted when in place in order to focus the air cooled laserrelative to a microplate bottom that the air cooled laser is shining on.By mounting the holder on a support the air cooled laser can be adjustedin three dimensional space relative to the microplate bottom surface.One can easily fashion such adjustment means within the scope of theinvention based on the disclosure herein and such is well within theskill in the art. The air cooled laser can be fixedly attached to suchsupport or they can be set by gravity or other means on the support asdesired. The fixed method of attaching to a support does allow thesupport and air cooled laser to be placed as a unit within the readerand thus makes replacement and set up much quicker and simpler.

In one embodiment, the air cooled laser is further contained within acase such as a box or the like. Such case would have a single opening ina single direction. Thus, stray light for the air cooled laser would beminimized and result in less overall light “noise” in reading theresults of experiments with the air cooled laser based reader. Sincescattered light has been discovered to be a potential problem with thepresent invention, where such is a problem additional dark, black or thelike material can be positioned around the inside of the reader asdesired to reduce the bounce light effect from the air cooled laser ofthe invention.

In order to change the light wavelength to the wavelength the readerneeds, or more particularly the cells within a microplate need, tofluoresce. One may need to position a light filter which changes thewavelength of the air cooled laser emissions to the desired wavelength.Where one is attempting to duplicate the argon water cooled laser afilter which will cause the wavelength to change to 488 nanometers wouldbe used unless as the example proposed the laser is already at the samewavelength. Typical pass filters are in the 480 to 550 nanometer rangebut can easily be chosen by one skilled in the art based on theselection of the air cooled laser and the desired end wavelength in viewof this disclosure. Other wavelengths as desired could be obtained withappropriate filters. Such filters are available in the art. Theluminosity is always decreased when passing through a filter and suchmust be considered and the luminosity matched to the particular filtersince each filter may filter out more or less of the luminosity of theoriginal air cooled laser lights. The size of the filter is sufficientto allow the air cooled laser light to pass entirely through the filter.Restricting the size of the filter can be used to focus and homogenizethe emitted light as desired and the size of the desired filter and thedistance from the air cooled laser while critical to the practice of theinvention can be determined with minimum experimentation by one skilledin the art in view of this disclosure. One embodiment has the filterattached to the air cooled laser case or in another embodiment attachedto or supported by the support that supports the air cooled laserlights.

The air cooled laser light must when replacing the water cooled laser,shine such that the filtered or unfiltered light shines on the bottomsurface of a microplate placed in the reader. This can be done from anangle (less than 90 degrees) as taught by the art cited above or inother embodiments shining at 90 degrees directly or by way of a bouncemirror. The bounce mirror, in some embodiments, can be a dichroic mirrorwhich allows the air cooled laser light to bounce off the dichroicmirror and hit the bottom surface of the microplate reader. Then whenthe emitted light comes from the bottom surface of the microplate, itcan pass down through the dichroic mirror to a receiver below the mirroras long as the angles are correct for use of such a mirror. In oneembodiment, a dichroic mirror that bounces light at 45 degrees is used.Air cooled laser light hits the mirror at 45 degrees and is reflected tothe bottom surface of a microplate in the reader. The emitted light fromthe microplate exits at 90 degrees passes through the dichroic mirrorand to the CCD camera in the reader positioned below the dichroicmirror. This can be seen as discussed with the figures which follow.

The support for the air cooled laser and optionally the filter ordichroic mirror serves at least two purposes. First, it keeps the partsin relative position to one another and makes 3 dimensional adjustmentseasier without changing their relative positions. Second, the supportallows the system to be placed in an individual reader in such a waythat it is immediately positioned for use by positioning the support inthe predetermined position for replacing the original water cooledmirror. This forgoes the problems of finding the optimum position foreach component and then doing the final 3D micro adjustments necessaryto align and focus the light. The complete support can be placed intothe reader and attached thereto and the installation is essentiallycomplete with only final adjustment calibrations necessary. Thisembodiment, either with or without the dichroic filter embodiment,allows for quick easy repair, easy adjustment and a lower cost to repairthan either replacing the laser or starting with each individualcomponent.

Lastly, it has been discovered that this system can be turned on and offas the laser was using essentially the same means within the reader.Such is surprising since the units were not designed to accept anythingother than water cooled argon laser light sources. The software ifnecessary can be modified as well but one skilled in the art can easilyprogram the unit consistant with the repair/replacement of the watercooled unit.

Now referring to the drawings, FIG. 1 is a side view of a method and aircooled laser system 1 of the present invention showing an air cooledlaser 2. The air cooled laser 2 is positioned partly outside themicroplate reader 3. In this example is shown the inside front view ofmicroplate reader 3. The upper interior 4 of the reader is where amicroplate 5 in which are positioned a number of wells 5 a. Microplate 5sits or rests on microplate support 6 and exposes the bottom surface 7of microplate 5 to lower interior 8. The laser light from the originalwater cooled laser in the original microplate reader shines on thebottom 7 and the excitation light from wells 5 a shines down from themicroplate 5 and is captured by a camera not shown in this view. In thisview a bounce mirror 10 is shown. The system 1 of the present inventionconsists of the air cooled laser 2 which is placed and attached tosupport 15 and optional holders to fix the laser 2 in place and aid withadjustment and focusing. The support 15 gives the air cooled laser 2 thecorrect height and position subject to final micro adjustments andallows the air cooled laser 2 to be placed as a unit without need fortotal customization for each unit. The laser 2 could also be positionedwithin a box, not shown, in order to reduce stray light and furtherfocus the light. Such box could be opaque and a dark or black color canbe used to further aid in preventing stray light. If a filter isnecessary, it can be placed in front of the laser opening 16 where laserbeam 17 exits from the laser 2. A filter holder or aperture device canbe attached to a box the reader 3 or mounted in any convenient manner ifpresent and could easily be accomplished in view of the disclosureherein. Note the holders and supports can be provided with means toadjust in 6 degrees of freedom the actual aiming of the laser 2 to focuson the bottom 7 of the microplate 5.

One tremendous advantage of the present system 1 is that the entire unitcan easily replace the laser of the unit by placement of the system inthe open compartment of the lower chamber 8 or as shown in themicroplate reader 3 as a single unit. It has been discovered that whenthe air cooled laser 2 is connected via wire 21 to the controls (notshown) of microplate reader 3, that a light system which does not haveall the frailty of the previous water cooled laser light source iscreated in a device originally designed for use only with a water cooledargon laser (3W). It is also determined that such can be accomplishedwithout the need to rebuild the reader 3 and mere refurbishment ispossible. One will note that the original mirror 10 can remain in placeand as such the camera which reads fluorescence from the microplate 5does not need to be adjusted or removed. Within the space given one canfit the system 1 of the unit in place and either have the componentspre-adjusted or individually adjust the laser 2 position and optionalfilter position relative to one another. By providing a support 15, itis possible to eliminate the need to constantly adjust the height of thesystem 1 relative to the bottom 7.

FIG. 2 involves a separate embodiment of the present invention where itis desirous to shine the laser 2 light at 90 degrees and where themirror would otherwise be in the way of the CCD camera. In thisembodiment, the system 1 consists of support 25 which supports the aircooled laser 2 and allows the entire system to be positioned inside thereader 3 in the lower interior 8. In order to utilize a 90 degree lightin this embodiment, a dichroic mirror 30 is used. The dichroic mirror 30is positioned at a 45 degree angle or as needed for the particularinstallation. Light shining from the laser 2 shines through an optionalfilter not shown and hits mirror 30 reflecting up and hitting bottom 7at 90 degrees. The emissions 18 generated in the wells 5 a shinesdownward at 90 degrees but passes directly through the dichroic filter30 and can hit original reflective mirror 10 (not shown in this figure)or picked up by CCD camera 31. This arrangement has the advantage ofonce again positioning the system in the lower interior 8 a direct beamand not a 45 degree angled beam. Non-reflective material can also beadded to interior 8 if reflectance becomes a problem with a particularrepair.

Air cooled laser replacement using the method and system of the presentinvention will add longevity to the FLIPR readers and has beendiscovered to increase the time between light related refurbishmentsince when using the system of the present invention refurbishment timesbetween work increases dramatically. In addition, a much lower powerconsumption can be used with the air cooled lasers, thus, saving energywhich could eventually lead to a cost saving that pays for a replacementeven when the water cooled laser does not need replacement. Accordingly,the results achieved with the present invention not only provide a quickand easy way to refurbish a FLIPR machine they surprisingly addlongevity to the readers and provide a result not anticipated with aircooled laser use in a refurbishment situation.

Nothing in the embodiments is designed to be limiting unless otherwisestated. In view of the disclosure of the embodiments once can easily seeif skilled in the art other substitutions of filters, support materials,air cooled lasers, and the like within the scope of the presentinvention. The claims which follow should not therefore be read as solimiting.

What is claimed is:
 1. A method for repairing a fluorescence microplatereader having a water cooled laser light source of about 3 watts thathas been removed from the reader comprising: a) selecting an air cooledlaser having a wattage of between about 300 and 500 mW, that produces areplacement lumen output, the air cooled laser mounted in a rigidsupport and provide light to the microplate reader; b) providing thatthe light output of the air cooled laser is at a desired wavelength; c)positioning the support such that the light from the air cooled lasershines on the bottom of a microplate positioned in the reader; and d)connecting the air cooled laser to the reader such that they operate inplace of the water cooled laser light.
 2. A method according to claim 1wherein the air cooled laser emits light in a wavelength from about 425to about 510 nanometers.